Differentiation of gastrointestinal endocrine cells depends on the expression and function of basic helix loop helix (bHLH) transcription factors. Notch signaling restricts the number of cells able to adopt an endocrine cell fate in the intestine, stomach, and pancreas by inhibiting the function of bHLH proteins. The overall goal of the proposed research is to determine at what stage of differentiation endocrine progenitor cells can revert to non endocrine cell fates, to determine when in development progenitor cells remain sensitive to the inhibitory effects of notch signaling, and to identify genes activated at different stages of enteroendocrine cell differentiation. Three specific aims are planned. The first aim will use a Cre/Iox based approach in transgenic mice to examine the cell fate of gastrointestinal endocrine precursor cells expressing bHLH proteins. The second aim will determine the developmental context when GI endocrine progenitors are regulated by notch and its associated protein, Deltex-1. For this aim, an activated form of notch 1 will be conditionally expressed at different stages of endocrine differentiation in transgenic mice to determine the stage when precursor cell differentiation can be inhibited by notch signals. The effects of Deltex-1 will also be examined in transgenic mice to determine whether this protein functions as a potentiator or antagonist of notch in the gastrointestinal tract. The third aim will identify genes differentially expressed in cells of the small intestine as they differentiate from endocrine precursors in crypts to terminally differentiated enteroendocrine cells in the villi. Enteroendocrine precursors at different developmental stages will be collected by laser microdissection for RNA extraction and preparation of probes to interrogate high-density microarrays. It is anticipated that these studies will provide important new insights regarding cell fate determination of GI endocrine cells, the mechanism of notch signaling in regulating endocrine differentiation in the GI tract, and genes that may regulate endodermal endocrine differentiation. Understanding how precursor cells become committed to differentiate into gastrointestinal endocrine cells may have an impact on potential new therapies for treating diseases like diabetes mellitus. Information from this project may contribute to the development of future treatments that involve manipulating stem cells or inducing other organs to transdifferentiate into cells capable of producing insulin or other hormones.